Hypoxic tumor cell radiosensitizers are compounds that selectively increase the sensitivity of hypoxic cells in tumors to radiation. This specificity for hypoxic cells is important because it is tumors that are typically characterized by such cells. All tumors which are present as solid masses contain these cells, while normal cells generally have an adequate supply of oxygen. The effects of radiation are enhanced by the presence of oxygen, and it is established that as the dose of radiation is increased, the effectiveness of the radiation in destroying target cells is enhanced most dramatically when oxygen is present.
Of course, the use of radiation treatment to destroy tumor cells is only practical if damage to the surrounding normal tissue can be minimized or avoided. The discussion in the foregoing paragraph demonstrates that this is difficult to achieve--normal cells, in view of their oxygen supply, are generally more susceptible to radiation than the target tumor cells. It is therefore desirable to provide a means to sensitize tumor cells, but not the surrounding tissue, to radiation treatment. One mechanism which would permit this is the augmentation of these cells' oxygen supply.
Various heterocyclic compounds, in particular, those with oxidized nitrogen moieties, have been used for the purpose of radiosensitizing tumor cells. Indeed, it has been postulated that the oxidized nitrogen functionality is responsible for this activity. Nitroimidazoles, particularly misonidazole (MIS) and metronidazole have been studied extensively, and MIS is commonly used as a standard in in vitro and in vivo tests for radiosensitizing activity. (See, e.g., Asquith, et al, Radiation Res (1974) 60:108-118; Hall, et al, Brit J Cancer (1978) 37: 567-569; Brown, et al, Radiation Res (1980) 82:171-190; and U.S. Pat. No. 4,371,540. The radiosensitizing activities of certain 1-substituted 3(5)-nitro-s-triazoles and of various quinoxaline-1,4-dioxide derivatives have also been disclosed.
The present invention provides a new group of radiosensitizers that do not contain oxidized nitrogen--the substituted benzamides and nicotinamides and their thio analogs. In this regard, it is important to distinguish the ability to sensitize hypoxic cells selectively, for instance, by enhancing their oxygen supply from another mechanism commonly encountered for "sensitizing" cells--inhibiting the enzyme poly(ADP-ribose)polymerase, which is believed to be essential in the repair of irradiated cells after radiation. This repair mechanism is operative in both hypoxic tumor cells and in normal cells. Hence, administration of "radiosensitizers" which exert their effects by this mechanism does not accomplish the desired purpose of selectively sensitizing the target tumor cells. While lower doses of radiation may be required to kill the tumor, normal cells, too, will experience a higher damage level at this lower dosage. This mechanism, which is called "potential lethal damage repair" (PLDR) can be distinguished experimentally from the desired selective radiosensitization by examining the kinetics of the effect. Since the repair occurs after the radiation is completed, agents which inhibit PLDR must be present during the repair period, and their presence during the irradiation itself is not required. On the other hand, true radiosensitizers must be present during the radiation treatment, and can be removed immediately thereafter, without change in their effect.
Ben Hur, E., et al, Radiation Res (1984) 97: 546-555, showed the effect of certain benzamide and nicotinamide derivatives on the repair mechanism in cell culture. Cultures of Chinese hamster ovary cells were used, and the effect of post-irradiation addition of these derivatives on cell survival was studied. It was found that several of these compounds indeed inhibited the repair mechanism, which the authors conclude is therefore relevant to enhancing the killing of those cells by radiation. Miyakoshi, J., et al. Radiation Res (1985) 102: 359-366, studied the effects of m-aminobenzamide on the survival, after radiation or hyperthermia, for Chinese hamster cells in culture. The m-aminobenzamide was added either during or after irradiation, and was found to sensitize these cells to the lethal effects of radiation. These cells are not hypoxic, since they are maintained in culture. The Miyakoshi et al results showed that the greatest enhancement ratio was obtained when cells were first treated with X-rays, then at increased temperature in the presence of m-aminobenzamide, followed by prolonged treatment with m-aminobenzamide. These authors also conclude that m-aminobenzamide exerts its effect by interference with the repair mechanism presumably present in all cells.
In vivo studies using nicotinamide were reported by Jonsson, G. G., et al, Cancer Res (1985) 45: 3609-3614. In these studies, the repair mechanism was also implicated in the effect of this compound in sensitizing the cells, even though the nicotinamide was administered prior to irradiating. The results obtained by Jonsson et al indicated that the enhancement ratio for the nicotinamide was greater for normal cells in vivo than that exhibited for tumor cells. It is believed that these results were obtained because cells in the normal cell model chosen (tail growth) are atypical of normal cells with regard to their oxygen supply. The authors, however, theorize that treatment with nicotinamide may sensitize cells both as a direct precursor for NAD.sup.+ synthesis and as an inhibitor to the repair mechanism. Based on this speculation, the authors suggest that nicotinamide may be useful in sensitizing tumor cells, despite their results to the contrary. Two additional reports on radiosensitization by nicotinamide of tumor cells are those of Jonsson, G. G., et al, Radiother and Oncol (1984) 1: 349-353, and of Calcutt, G., et al, Br J Cancer (1970) 24: 380-388.
In the invention herein, both benzamide and nicotinamide derivatives and their thio analogs have been shown to have selective radiosensitizing activity in vivo. This effect has been shown to be due to enhancement of oxygen supply and to exhibit a time dependence which corresponds to direct radiosensitazation. Therefore, administration of these compounds prior to radiation is sensitizing selectively for hypoxic (tumor) cells. This is an unexpected property even for those compounds which are known to inhibit poly(ADP-ribose)polymerase.